1. Field of the Invention
This invention relates to identification of components in a biological or chemical sample by optical trapping means.
2. Description of Related Art
Numerous instruments, kits and methods generally related to the identification and quantitation of chemical and biochemical moieties are currently available. At present, none of these rely on an "optical trapping means" or "optical tweezers" as described herein, to manipulate the reaction substrate for the purpose of controlling the reaction, and for the purpose of obtaining accurate measurement using only a small sample size. The optical trapping means consists essentially of a laser capable of emitting a beam of suitable wavelength (e.g., Nd:YAG laser). The beam impinges upon a microparticle (e.g., a 5 micron polystyrene bead which serves as a reaction substrate), and the bead is thus confined at the focus of the laser beam by a radial component of the gradient force. Once "trapped" the bead can be moved, either by moving the beam focus, or by moving the reaction chamber. In this manner, the bead can be transferred among separate reaction wells, to permit reaction between the reagent affixed to the bead, and the reagent contained in that well.
This optical trapping means has been previously characterized in the art and is the subject of U.S. Pat. No. 4,893,886 to Ashkin, Non-destructive optical trapping means for biological particles and method of doing same, assigned to AT&T, and issued on Jan. 16, 1990.
The apparatus, method, and system of the present invention is suitable for a variety of applications: chemistry, biochemistry, immunology, etc. Applications for which the present invention are presently suited are immunological assays ("immunoassay"). Such techniques are directed to, for example, probing antigen-antibody interactions.
Briefly, an antibody is a molecule produced by the immune system of an animal in response to a foreign particle or pathogen (e.g., a disease-causing bacterium). The antibody is able to recognize (chemically bond to) a particular portion of the foreign particle known as the antigen; a single foreign particle may have several antigens, though a particular antibody binds to only one of them. This recognition and subsequent binding are among the initial stages in the immune response. Hence, specific antibodies are produced by the body in response to particular pathogens. And therefore, the presence of a particular antibody in the blood is a reliable indicator of a particular infection, which may be found long before the onset of any signs or symptoms of the disease. Indeed, since at a given moment, antibody levels may far exceed the pathogen levels in the body, it is far easier to look for the presence of the antibody--as an indicator of the disease--than it is to look for the pathogen directly. Consequently, one common technique for determining whether a person is infected with a disease-causing pathogen is to assay for the presence of particular antibodies.
In addition, antibodies form very strong chemical bonds with a particular antigen found on the surface of the pathogen. Thus, particular antigens can be isolated, and used as "probes," in the following manner. A small sample of blood is taken from a person; the antigen probe is then added to the sample. If the blood sample contains antibody specific for that antigen, then it will chemically bind to the antigen probe. Finally, the investigator needs a means to determine whether this binding has occurred. This can be done in a variety of ways. Perhaps the simplest way relies upon the fact that an antigen-antibody complex will precipitate out of solution (i.e., form a solid and settle out of the solution) allowing it to be easily identified, even by visual inspection. Again, numerous immunological techniques are currently available either commercially or described in the scientific literature. A concise review of these techniques is provided in Immunolog, Roitt, et al. eds. (1996), which is hereby incorporated by reference into the present Application. The simplest such method--just discussed--is probably the precipitin reaction, which exploits the tendency of antigen-antibody complexes to precipitate from solution when combined in proportions at or near equivalence. Hemagglutination and complement fixation describe more sophisticated techniques, which allow antibody to be detected and measured at far lower concentrations than those detectable by the method just discussed; these methods can readily detect antibody at levels of less than 1 .mu.g/ml. Direct and indirect immunofluorescence relies upon fluoresceinated antibody which, when exposed to UV light, will fluoresce green if bound with antigen. Immunoassay techniques rely on labeled reagents (e.g., fluorescent, chemiluminescent, or radioisotope markers) for detecting antigens and antibodies, and include both solid-phase assays (one reagent fixed to a solid support) and ELISA (enzyme-linked immunoabsorbent assay). One such system is described in U.S. Pat. No. 4,623,629, Solid-phase immunoassay support and method of use thereof, issued to Daniel Kerschensteiner. Magnetic beads can be moved by forces of a magnetic field, these forces typically are not confined to a single bead.
Current analytical techniques are generally plagued by the following problems: (1) the sample size is too small to reliably identify the sought-after substance; and (2) the technique is too slow and/or is not amenable to automation. The apparatus and method of the present invention addresses these problems in the following way. One, the reaction substrate is a single micron-sized bead; moreover, the detection method is in constant coincidence with the bead location, which is fixed by the optical trapping means, hence the measurement is highly sensitive and easily automated. Second, because the bead is readily transferred from one reservoir to another, measurement can take place in a "clean" reservoir, free of background reagents, which allows more accurate measurement. Finally, random access to bead(s) on which different chemical are coated, can enable multianalyte analysis.